Isolation and characterization of hematopoietic stem cells from human umbilical cord blood for cell therapy
Author(s): Durga Prameela G., T.N.V.K.V.Prasad*, M.Nagalakshmi Devamma, E.K.Elumalai, P.M. Ayyasamy and T.Kiran Reddy
Abstract
Alternatives to organ transplantation for treatment of certain liver diseases are currently under investigation. Whole liver, split liver and related living donor liver transplantation are clinically safe and well established procedures for the treatment of end-stage liver failure. However, organ donor shortage and the need of lifelong immunosuppressive treatment are still the major limitations of all these options. Therefore, the development of cell based therapeutic strategies for liver disease is under experimental evaluation. Umbilical cord blood was collected and isolated the mononucleated cell by the help of ficoll hypaque centrifugation. The collected samples were purified for CD 34+ hematopoietic stem cells using MACS. These purified hematopoietic stem cells were counted and morphologically characterized by FACS. Freshly isolated CD 34+ cells were cultured in hepatocyte differentiation media; contain growth factor of HGF and FGF-4. These growth factors induced the differentiation of hematopoietic stem cells into hepatocyte like cells. The cell density played a significant role in effecting hepatic differentiation. Optimal cell density was found to be 1.0 to 1.3X10â´ cells /cm². After 14 inductions, cuboidal morphology, which is characteristic of hepatocytes were observed and cells also expressed by marker genes specific for liver cells in a time depended manner.
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